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Rational diversification of a promoter providing fine-tuned expression and orthogonal regulation for synthetic biology.
Blount, Benjamin A; Weenink, Tim; Vasylechko, Serge; Ellis, Tom.
Afiliação
  • Blount BA; Centre for Synthetic Biology and Innovation, Imperial College London, London, United Kingdom.
PLoS One ; 7(3): e33279, 2012.
Article em En | MEDLINE | ID: mdl-22442681
ABSTRACT
Yeast is an ideal organism for the development and application of synthetic biology, yet there remain relatively few well-characterised biological parts suitable for precise engineering of this chassis. In order to address this current need, we present here a strategy that takes a single biological part, a promoter, and re-engineers it to produce a fine-graded output range promoter library and new regulated promoters desirable for orthogonal synthetic biology applications. A highly constitutive Saccharomyces cerevisiae promoter, PFY1p, was identified by bioinformatic approaches, characterised in vivo and diversified at its core sequence to create a 36-member promoter library. TetR regulation was introduced into PFY1p to create a synthetic inducible promoter (iPFY1p) that functions in an inverter device. Orthogonal and scalable regulation of synthetic promoters was then demonstrated for the first time using customisable Transcription Activator-Like Effectors (TALEs) modified and designed to act as orthogonal repressors for specific PFY1-based promoters. The ability to diversify a promoter at its core sequences and then independently target Transcription Activator-Like Orthogonal Repressors (TALORs) to virtually any of these sequences shows great promise toward the design and construction of future synthetic gene networks that encode complex "multi-wire" logic functions.
Assuntos

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Saccharomyces cerevisiae / Regulação Fúngica da Expressão Gênica / Regiões Promotoras Genéticas / Biologia Sintética Idioma: En Revista: PLoS One Assunto da revista: CIENCIA / MEDICINA Ano de publicação: 2012 Tipo de documento: Article País de afiliação: Reino Unido

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Saccharomyces cerevisiae / Regulação Fúngica da Expressão Gênica / Regiões Promotoras Genéticas / Biologia Sintética Idioma: En Revista: PLoS One Assunto da revista: CIENCIA / MEDICINA Ano de publicação: 2012 Tipo de documento: Article País de afiliação: Reino Unido